The lack of effective treatments for many diseases underscores the need to discover new drugs. Natural products, which are chemical compounds produced by organisms, have had a rich history of being used as pharmaceutical drugs. However, diminishing returns of new natural products from traditional sources, namely actinomycete bacteria, compounded with the problem of rediscovery necessitates finding them elsewhere. This proposal involves prospecting for natural products from an unconventional source - bacteria in symbiosis with insects in eusocial agricultural systems, and using the distribution and evolutionary history data for these discovered natural products to find analogs in other systems. The proposed project will be carried out in Prof. Jon Clardy's laboratory in the Department of Biological Chemistry and Molecular Pharmacology at Harvard Medical School. In regards to the overall strategy, the project can be split into three objectives. The first is to identify and prioritize bacterial symbionts from insect agricultural systems that produce novel natural products. The expanding insect-associated actinobacteria collection in collaborator Prof. Cameron Currie's lab will be subjected to next generation sequencing to form draft genomes. These will be queried with biosynthetic gene clusters for natural products with known activity to find analogs, and scanned thoroughly using bioinformatics software developed in collaborator Prof. Michael Fischbach's laboratory to unearth many other biosynthetic clusters, including those that are cryptic and/or very novel. The second specific aim involves the isolation and characterization of new natural products from the prioritized set of bacterial symbionts. This involves growing bacteria under various conditions to elicit expression of constitutively-expressed and cryptic natural products, and identifying their production using differential metabolomics or bioassay-guided fractionation. The structures of the isolated natural products will be solved, and therapeutic potential will be characterized through assays in the Clardy lab and in conjunction with the Institute of Chemistry and Cell Biology - Longwood at Harvard Medical School. The third aim is ascertaining the evolutionary history and distribution of the novel bioactive natural products. Bioinformatics methods will be used to link the natural products back to the biosynthetic gene cluster. These clusters will be compared to others in the span of actinobacteria, giving insight into distribution within the insect-associated actinobacteria niche and beyond. Evolutionary studies of the clusters will be performed through genetic synteny and phylogenetic analysis. The distribution and evolutionary data will allow for identification of analogs from other actinobacteria and other bacterial phyla. The evolutionary data will especially help in tracking down markedly different analogs.